: Despite the fact that low postischemic adenosine triphosphate (ATP) levels are correlated with postoperative ventricular function of failure in infants, information regarding ATP metabolism in immature hearts is scarce. The overall goals of this project is to clearly define the biochemical pathways associated with adenine nucleotide metabolism in neonatal piglet myocytes, to characterize the effect of simulated ischemia on energy producing cellular processes and ATP catabolic pathways, and to evaluate the ability of cardiac myocytes to restore ATP during reperfusion. Differences between neonatal and adult ATP generating and degradative routes will be delineated and differences between the neonate and adult in response to several therapeutic interventions which are cardioprotective in adult hearts will be examined. For these studies cardiac myocytes will be isolated from new born pig hearts during a collagenase disaggregation procedure. Ventricular myocytes are then incubated under conditions designed to simulate in vivo ischemia and reperfusion. Complete nucleotide and nucleoside profiles of cells will be obtained using HPLC. Changes in phosphocreatine and total creatine, lactate production (a measure of glycolytic activity) and respiration rates (an estimate of mitochondrial competence) will be analyzed as well. In selected protocols, the effect of pharmacological preconditioning on electrically stimulated single cell calcium transient and extent of cell shortening will be monitored. The specific aims of the project are (i) to fully characterize patterns of ATP degradation and synthesis during simulated ischemia and reperfusion in the piglet myocytes, and (ii) to examine the effects of adenosine, alpha 1- adrenergic, muscarinic or cholinergic stimulation, and preconditioning on ATP metabolic pathways during ischemia-reperfusion. It is postulated that a metabolic differences in the neonate alter the overall cellular response to ischemia and reperfusion. Differences between the neonate and adult in glycolytic capacity and in the enzymatic machinery responsible for purine metabolism may contribute to these alterations. The hypothesis for Specific Aim two is that mechanisms which promote degradation of AMP to inosine monophosphate (IMP) rather than to adenosine will enhance postischemic restoration of ATP. The alternative hypothesis is that stimulation of this pathway is detrimental and may be a crucial element in the inability of neonates to tolerate prolong ischemia. These data may assist in the development of therapeutic interventions specifically designed to protect the globally ischemic, immature heart.